Curcuphenol compounds for increasing mhc-i expression

ABSTRACT

Provided are methods of using curcuphenol compounds to increase expression of major histocompatibility complex class I (MHC-I) antigen in cells, particularly on the surface of diseased cells such as cancer cells, and thereby increase the immunogenicity of the cells. Also provided are pharmaceutical compositions that comprise curcuphenol compounds and methods of use thereof, for instance, to treat various cancers, alone or in combination with other therapies.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a Continuation of U.S. application Ser. No.14/548,726, filed Nov. 20, 2014; which claims priority under 35 U.S.C.119(e) to U.S. Application No. 61/906,817, filed Nov. 20, 2013, which isincorporated by reference in its entirety.

BACKGROUND Technical Field

Embodiments of the present invention relate to the use of curcuphenolcompounds for increasing expression of major histocompatibility complexclass I (MHC-I) antigen in cells, particularly on the surface ofdiseased cells such as cancer cells, and thereby increasing theimmunogenicity of the cells. Also included are related pharmaceuticalcompositions and methods of use thereof, for instance, to treat variouscancers, alone or in combination with other therapies.

Description of the Related Art

Major histocompatibility complex class I (MHC-I) antigens are found onnearly all nucleated cells of the body. The primary function of thisclass of major histocompatibility complex (MHC) molecules is to display(or present) peptide fragments of intracellular proteins to cytotoxic Tlymphocytes (CTLs). Based on this display, CTLs ignore will healthycells and attack those displaying MHC-bound foreign or otherwiseabnormal peptides, including disease-associated peptide (antigens) suchas cancer antigens. Thus, the surface expression of MHC-I moleculesplays a crucial role in determining the susceptibility of target cellsto CTLs. Many cancerous cells display down-regulated MHC-I cell surfaceexpression (see, for example, Wang et al., JBC. 283: 3951-3959, 2008;Chang et al., Keio J. Med. 52:220-9, 2003; Zagzag et al., Lab Invest.85:328-41, 2005; and Hewitt, Immunology. 110:163-69, 2003). ReducedMHC-I expression can result at least in part from the down-regulation ofmultiple factors such as transporters (for example, TAP-1, TAP-2),proteasome components (LMP), and other accessory proteins involved inthe antigen presentation and processing pathway. This characteristic mayallow cancerous cells to evade immune surveillance and thereby provide asurvival advantage against immune activity otherwise designed toeliminate the cells.

Accordingly, there is a need in the art for agents that can increase MHCclass I expression in these and other types of diseased cells andthereby improve the ability of the immune system to target such cellsfor destruction.

BRIEF SUMMARY OF THE INVENTION

Embodiments of the present invention include methods for increasingmajor histocompatibility complex class I (MHC-I) surface expression in acell, comprising contacting the cell with a curcuphenol compound. Incertain embodiments, MHC-I surface expression is increased by at leastabout 10% relative to an untreated control cell.

In certain embodiments, the cell (in its untreated state) ischaracterized by reduced MHC-I surface expression and optionally reducedTAP-1 expression relative to a normal or otherwise healthy cell of thesame cell type. In certain embodiments, MHC-I surface expression andoptionally TAP-1 expression in the cell is increased to within about 10%of the levels of MHC-I surface expression and optionally TAP-1expression of the otherwise normal or healthy cell of the same celltype.

In certain embodiments, the cell is a cancer cell. In certainembodiments, the cancer cell is a metastatic cancer cell. In certainembodiments, the cancer cell is selected from one or more of a breastcancer cell, a cervical cancer cell, a prostate cancer cell, agastrointestinal cancer cell, a lung cancer cell, an ovarian cancercell, a testicular cancer cell, a head and neck cancer cell, a bladdercancer cell, a kidney cancer cell, a squamous cell carcinoma, a CNS orbrain cancer cell, a melanoma cell, a non-melanoma cancer cell, athyroid cancer cell, a endometrial cancer cell, an epithelial tumorcell, a bone cancer cell, and a hematopoietic cancer cell. In certainembodiments, the bone cancer cell is an osteosarcoma, chondrosarcoma, ora cell of the Ewing Sarcoma Family of Tumors (ESFTs). In certainembodiments, the gastrointestinal cancer cell is an esophageal cancercell, stomach (gastric) cancer cell, pancreatic cancer cell, livercancer cell, gallbladder (biliary) cancer cell, small intestinal cancercell, colorectal cancer cell, anal or rectal cancer cell, or agastrointestinal carcinoid or stromal tumor. In certain embodiments, thelung cancer cell is an adenocarcinoma, squamous-cell lung carcinoma,small-cell lung carcinoma, or a large-cell lung carcinoma. In certainembodiments, the melanoma is a lentigo maligna, lentigo malignamelanoma, superficial spreading melanoma, acral lentiginous melanoma,mucosal melanomas, nodular melanoma, polypoid melanoma, desmoplasticmelanoma, amelanotic melanoma, soft-tissue melanoma, or a uvealmelanoma. In certain embodiments, the hematopoietic cancer cell is alymphoma cell, leukemia cell, or a multiple myeloma cell.

In certain embodiments, the cell is in vitro.

In certain embodiments, the cell is in a subject, and the methodcomprises administering the curcuphenol compound to the subject. Incertain embodiments, the subject has cancer. In certain embodiments, thecancer is characterized by cancer cells (in an untreated state) havingreduced MHC-I surface expression and optionally reduced TAP-1 expressionrelative to non-cancerous cells of the same cell type. In certainembodiments, the cancer cell(s) comprise metastatic cancer cells. Incertain embodiments, the metastatic cancer cells (in an untreated state)have reduced MHC-I surface expression and optionally reduced TAP-1expression relative to non-cancerous cells of the same cell type, orrelative to non-metastatic cancer cells of the same cell type.

In certain embodiments, MHC-I surface expression and optionally TAP-1expression in the cancer cell(s) is increased by at least about 10%relative to a control cell. In certain embodiments, increased MHC-Isurface expression and optionally TAP-1 expression increases aCTL-mediated immune response against the cancer cells.

In certain embodiments, the cancer is selected from one or more ofbreast cancer, cervical cancer, prostate cancer, gastrointestinalcancer, lung cancer, ovarian cancer, testicular cancer, head and neckcancer, bladder cancer, kidney cancer (e.g., renal cell carcinoma), softtissue sarcoma, squamous cell carcinoma, CNS or brain cancer, melanoma,non-melanoma cancer, thyroid cancer, endometrial cancer, an epithelialtumor, bone cancer, and hematopoietic cancer. In certain embodiments,the lung cancer is osteosarcoma, chondrosarcoma, or a Ewing SarcomaFamily of Tumors (ESFTs). In certain embodiments, the gastrointestinalcancer is esophageal cancer, stomach (gastric) cancer, pancreaticcancer, liver cancer, gallbladder (biliary) cancer, small intestinalcancer, colorectal cancer, anal or rectal cancer, or gastrointestinalcarcinoid or stromal tumor. In certain embodiments, the melanoma islentigo maligna, lentigo maligna melanoma, superficial spreadingmelanoma, acral lentiginous melanoma, mucosal melanoma, nodularmelanoma, polypoid melanoma, desmoplastic melanoma, amelanotic melanoma,soft-tissue melanoma, or uveal melanoma. In certain embodiments, thehematopoietic cancer is a lymphoma, leukemia, or multiple myeloma. Incertain embodiments, the lymphoma is a T-cell lymphoma, B-cell lymphoma,small lymphocytic lymphoma, mangle cell lymphoma, anaplastic large celllymphoma (ALCL), follicular lymphoma, Hodgkin's lymphoma, ornon-Hodgkin's lymphoma. In certain embodiments, the leukemia is chroniclymphocytic leukemia (CLL), hairy cell leukemia, acute lymphoblasticleukemia, myelocytic leukemia, acute myeloid or myelogenous leukemia, orchronic myelogenous leukemia. In certain embodiments, the brain canceris a glioma, meningioma, pituitary adenoma, vestibular schwannoma,primary CNS lymphoma, neuroblastoma, primitive neuroectodermal tumor(medulloblastoma), or glioblastoma multiforme.

Some methods include administering the curcuphenol compound incombination with an additional cancer therapy. In certain embodiments,the additional cancer therapy selected from one or more of ananti-cancer agent, radiotherapy, surgery, transplantation, photodynamictherapy, symptomatic care, and antibiotic therapy. In certainembodiments, the anti-cancer agent is selected from a small molecule andan antibody. In certain embodiments, the small molecule is a cytotoxic,chemotherapeutic, or anti-angiogenic agent. In certain embodiments, thesmall molecule cytotoxic, chemotherapeutic, or anti-angiogenic agent isselected from one or more of alkylating agents, anti-metabolites,anthracyclines, anti-tumor antibiotics, platinums, type I topoisomeraseinhibitors, type II topoisomerase inhibitors, vinca alkaloids, andtaxanes.

In certain embodiments, the small molecule is selected from one or moreof chlorambucil, cyclophosphamide, cilengitide, lomustine (CCNU),melphalan, procarbazine, thiotepa, carmustine (BCNU), enzastaurin,busulfan, daunorubicin, doxorubicin, gefitinib, erlotinib idarubicin,temozolomide, epirubicin, mitoxantrone, bleomycin, cisplatin,carboplatin, oxaliplatin, camptothecins, irinotecan, topotecan,amsacrine, etoposide, etoposide phosphate, teniposide, temsirolimus,everolimus, vincristine, vinblastine, vinorelbine, vindesine, CT52923,paclitaxel, imatinib, dasatinib, sorafenib, pazopanib, sunitnib,vatalanib, geftinib, erlotinib, AEE-788, dichoroacetate, tamoxifen,fasudil, SB-681323, semaxanib, donepizil, galantamine, memantine,rivastigmine, tacrine, rasigiline, naltrexone, lubiprostone, safinamide,istradefylline, pimavanserin, pitolisant, isradipine, pridopidine(ACR16), tetrabenazine, bexarotene, glatirimer acetate, fingolimod, andmitoxantrone, including pharmaceutically acceptable salts and acidsthereof.

In certain embodiments, the antibody is selected from one or more of3F8, 8H9, abagovomab, adecatumumab, afutuzumab, alacizumab (pegol),alemtuzumab, altumomab pentetate, amatuximab, anatumomab mafenotox,apolizumab, arcitumomab, bavituximab, bectumomab, belimumab,bevacizumab, bivatuzumab (mertansine), brentuximab vedotin, cantuzumab(mertansine), cantuzumab (ravtansine), capromab (pendetide), carlumab,catumaxomab, cetuximab, citatuzumab (bogatox), cixutumumab, clivatuzumab(tetraxetan), conatumumab, dacetuzumab, daclizumab, dalotuzumab,detumomab, drozitumab, ecromeximab, edrecolomab, elotuzumab,enavatuzumab, ensituximab, epratuzumab, ertumaxomab, etaracizumab,farletuzumab, FBTA05, figitumumab, flanvotumab, galiximab, gemtuzumab,ganitumab, gemtuzumab (ozogamicin), girentuximab, glembatumumab(vedotin), ibritumomab tiuxetan, icrucumab, igovomab, indatuximabravtansine, intetumumab, inotuzumab ozogamicin, ipilimumab (MDX-101),iratumumab, labetuzumab, lexatumumab, lintuzumab, lorvotuzumab(mertansine), lucatumumab, lumiliximab, mapatumumab, matuzumab,milatuzumab, mitumomab, mogamulizumab, moxetumomab (pasudotox),nacolomab (tafenatox), naptumomab (estafenatox), narnatumab,necitumumab, nimotuzumab, nivolumab, Neuradiab® (with or withoutradioactive iodine), NR-LU-10, ofatumumab, olaratumab, onartuzumab,oportuzumab (monatox), oregovomab, panitumumab, patritumab, pemtumomab,pertuzumab, pritumumab, racotumomab, radretumab, ramucirumab,rilotumumab, rituximab, robatumumab, samalizumab, sibrotuzumab,siltuximab, tabalumab, tanezumab, taplitumomab (paptox), tenatumomab,teprotumumab, TGN1412, ticilimumab, trastuzumab, tremelimumab,tigatuzumab, TNX-650, tositumomab, TRBS07, tucotuzumab (celmoleukin),ublituximab, urelumab, veltuzumab, volociximab, votumumab, andzalutumumab, including antigen-binding fragments thereof.

Also included are compositions for use in treating cancer, comprising apharmaceutically acceptable carrier and a curcuphenol compound orpharmaceutically-acceptable salt thereof. In certain embodiments, thecancer is characterized by cancer cells (in an untreated state) havingreduced MHC-I surface expression and optionally reduced TAP-1 expressionrelative to non-cancerous cells of the same cell type.

Some embodiments include compositions (e.g., pharmaceuticalcompositions), comprising a pharmaceutically acceptable carrier, ananti-cancer agent, and a curcuphenol compound orpharmaceutically-acceptable salt thereof.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 shows the formulae of (S)-(+)-curcuphenol and various othercurcuphenol compounds.

FIGS. 2A-2B show the formulae of curcuphenol compounds produced bycoupling of curcuphenol with various carboxylic acids.

FIG. 3 illustrates the recognition of tumor-associated antigens oncancer cells by cytotoxic T lymphocytes (CTLs).

FIG. 4A illustrates the MHC class I antigen presentation pathway,highlighting the role of TAP-1 in the formation of peptide/MHC-Icomplexes. FIG. 4B illustrates the various mechanisms by which cancercells reduce display of peptide/MHC-I complexes on the cell surface andthereby evade CTL-mediated immune surveillance, including thedown-regulation of TAP-1 and TAP-2 transporter proteins and proteasomecomplexes.

FIG. 5 shows that TAP-1 and MHC-I are frequently down-regulated incancer cell lines and surgically removed tumors.

FIG. 6 shows that TAP-1 down-regulation strongly correlates with diseaseprogression and metastasis.

FIGS. 7A-7B show that marine sponge extract 1 increased surfaceexpression of MHC-I (7A) and expression at the TAP-1 promoter (7B, asindicated by EGFP).

FIGS. 8A-8B show that marine sponge extract 3 increased surfaceexpression of MHC-I (8A) and expression at the TAP-1 promoter (8B, asindicated by EGFP).

FIGS. 9A-9B show that marine sponge extract 5 increased surfaceexpression of MHC-I (9A) and expression at the TAP-1 promoter (9B, asindicated by EGFP).

FIG. 10 shows the formulae of the curcuphenol compounds that were testedin the A9 murine tumor cell model (see Example 2).

FIG. 11A shows the induction of MHC-I expression in A9 cells bycurcuphenol compounds, and FIG. 11B shows the results of cell toxicitystudies in A9 cells, as measured by PI exclusion and intact cell number.

DETAILED DESCRIPTION OF THE INVENTION

Embodiments of the present invention relate to the discovery thatcurcuphenol compounds can increase MHC-I cell surface expression. Insome instances, curcuphenol compounds achieve this effect by increasingthe expression of TAP-1 (Transporter associated with Antigen Processing1), a transporter protein of the MHC-I antigen presentation pathway.

Recognition of MHC-1/peptide complexes is crucial for CTL-mediatedimmune surveillance of cells (see FIG. 3). Because certain diseasedcells such as cancerous cells evade immune surveillance bydown-regulating MHC-I cell surface expression, often by down-regulatingexpression proteins of the antigen presentation pathway such as TAP-1(see FIGS. 4A-4B and 5-6), curcuphenol compounds may improveCTL-mediated immune activity towards these diseased cells by restoringMHC-I surface expression and presentation of MHC-I/peptide antigencomplexes. Curcuphenol compounds may thus find utility in the treatmentof diseases associated with reduced MHC-I surface expression and/orTAP-1 expression, including many cancers.

Definitions

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood by those of ordinary skillin the art to which the invention belongs. Although any methods andmaterials similar or equivalent to those described herein can be used inthe practice or testing of the present invention, preferred methods andmaterials are described. For the purposes of the present invention, thefollowing terms are defined below.

All publications, patents, and patent applications cited herein areincorporated by reference in their entireties.

The articles “a” and “an” are used herein to refer to one or to morethan one (i.e., to at least one) of the grammatical object of thearticle. By way of example, “an element” means one element or more thanone element.

By “about” is meant a quantity, level, value, number, frequency,percentage, dimension, size, amount, weight or length that varies by asmuch as 30, 25, 20, 15, 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1% to a referencequantity, level, value, number, frequency, percentage, dimension, size,amount, weight or length.

Throughout this specification, unless the context requires otherwise,the words “comprise,” “comprises,” and “comprising” will be understoodto imply the inclusion of a stated step or element or group of steps orelements but not the exclusion of any other step or element or group ofsteps or elements. By “consisting of” is meant including, and limitedto, whatever follows the phrase “consisting of.” Thus, the phrase“consisting of” indicates that the listed elements are required ormandatory, and that no other elements may be present. By “consistingessentially of” is meant including any elements listed after the phrase,and limited to other elements that do not interfere with or contributeto the activity or action specified in the disclosure for the listedelements. Thus, the phrase “consisting essentially of” indicates thatthe listed elements are required or mandatory, but that other elementsare optional and may or may not be present depending upon whether or notthey materially affect the activity or action of the listed elements.

The terms “modulating” and “altering” include “increasing,” “enhancing”or “stimulating,” as well as “decreasing” or “reducing,” typically in astatistically significant or a physiologically significant amount ordegree relative to a control. An “increased,” “stimulated” or “enhanced”amount is typically a “statistically significant” amount, and mayinclude an increase that is 1.1, 1.2, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15,20, 30 or more times (e.g., 500, 1000 times) (including all integers anddecimal points and ranges in between and above 1, e.g., 1.5, 1.6, 1.7.1.8, etc.) the amount produced by no composition or a controlcomposition, sample, state, or test subject. A “decreased” or “reduced”amount is typically a “statistically significant” amount, and mayinclude a 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%,15%, 16%, 17%, 18%, 19%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%,65%, 70%, 75%, 80%, 85%, 90%, 95%, or 100% (including all integers andranges in between) decrease in the amount produced by no composition ora control composition, sample, state, or test subject.

“Optional” or “optionally” means that the subsequently described eventor circumstances may or may not occur, and that the description includesinstances where said event or circumstance occurs and instances in whichit does not.

“Pharmaceutically acceptable carrier, diluent or excipient” includeswithout limitation any adjuvant, carrier, excipient, glidant, sweeteningagent, diluent, preservative, dye/colorant, flavor enhancer, surfactant,wetting agent, dispersing agent, suspending agent, stabilizer, isotonicagent, solvent, or emulsifier which has been approved by the UnitedStates Food and Drug Administration as being acceptable for use inhumans or domestic animals.

“Pharmaceutically acceptable salt” includes both acid and base additionsalts.

“Pharmaceutically acceptable acid addition salt” refers to those saltswhich retain the biological effectiveness and properties of the freebases, which are not biologically or otherwise undesirable, and whichare formed with inorganic acids such as, but are not limited to,hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid,phosphoric acid and the like, and organic acids such as, but not limitedto, acetic acid, 2,2-dichloroacetic acid, adipic acid, alginic acid,ascorbic acid, aspartic acid, benzenesulfonic acid, benzoic acid,4-acetamidobenzoic acid, camphoric acid, camphor-10-sulfonic acid,capric acid, caproic acid, caprylic acid, carbonic acid, cinnamic acid,citric acid, cyclamic acid, dodecylsulfuric acid, ethane-1,2-disulfonicacid, ethanesulfonic acid, 2-hydroxyethanesulfonic acid, formic acid,fumaric acid, galactaric acid, gentisic acid, glucoheptonic acid,gluconic acid, glucuronic acid, glutamic acid, glutaric acid,2-oxo-glutaric acid, glycerophosphoric acid, glycolic acid, hippuricacid, isobutyric acid, lactic acid, lactobionic acid, lauric acid,maleic acid, malic acid, malonic acid, mandelic acid, methanesulfonicacid, mucic acid, naphthalene-1,5-disulfonic acid,naphthalene-2-sulfonic acid, 1-hydroxy-2-naphthoic acid, nicotinic acid,oleic acid, orotic acid, oxalic acid, palmitic acid, pamoic acid,propionic acid, pyroglutamic acid, pyruvic acid, salicylic acid,4-aminosalicylic acid, sebacic acid, stearic acid, succinic acid,tartaric acid, thiocyanic acid, p-toluenesulfonic acid, trifluoroaceticacid, undecylenic acid, and the like.

“Pharmaceutically acceptable base addition salt” refers to those saltswhich retain the biological effectiveness and properties of the freeacids, which are not biologically or otherwise undesirable. These saltsare prepared from addition of an inorganic base or an organic base tothe free acid. Salts derived from inorganic bases include, but are notlimited to, the sodium, potassium, lithium, ammonium, calcium,magnesium, iron, zinc, copper, manganese, aluminum salts and the like.Preferred inorganic salts are the ammonium, sodium, potassium, calcium,and magnesium salts. Salts derived from organic bases include, but arenot limited to, salts of primary, secondary, and tertiary amines,substituted amines including naturally occurring substituted amines,cyclic amines and basic ion exchange resins, such as ammonia,isopropylamine, trimethylamine, diethylamine, triethylamine,tripropylamine, diethanolamine, ethanolamine, deanol,2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine,lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline,betaine, benethamine, benzathine, ethylenediamine, glucosamine,methylglucamine, theobromine, triethanolamine, tromethamine, purines,piperazine, piperidine, N-ethylpiperidine, polyamine resins and thelike. Particularly preferred organic bases are isopropylamine,diethylamine, ethanolamine, trimethylamine, dicyclohexylamine, cholineand caffeine.

Often crystallizations produce a solvate of the curcuphenol compound. Asused herein, the term “solvate” refers to an aggregate that comprisesone or more molecules of a curcuphenol compound with one or moremolecules of solvent. The solvent may be water, in which case thesolvate may be a hydrate. Alternatively, the solvent may be an organicsolvent. Thus, the curcuphenol compounds may exist as a hydrate,including a monohydrate, dihydrate, hemihydrate, sesquihydrate,trihydrate, tetrahydrate and the like, as well as the correspondingsolvated forms. In some instances the curcuphenol compounds may be truesolvates, while in other instances the compounds may merely retainadventitious water or be a mixture of water plus some adventitioussolvent.

A “pharmaceutical composition” refers to a formulation of a curcuphenolcompound and a medium generally accepted in the art for the delivery ofthe biologically active compound to mammals, e.g., humans. Such a mediumincludes all pharmaceutically acceptable carriers, diluents orexcipients therefor.

The curcuphenol compounds, or their pharmaceutically acceptable saltsmay contain one or more asymmetric centers and may thus give rise toenantiomers, diastereomers, and other stereoisomeric forms that may bedefined, in terms of absolute stereochemistry, as (R)- or (S)— or, as(D)- or (L)- for amino acids. The present invention is meant to includeall such possible isomers, as well as their racemic and optically pureforms. Optically active (+) and (−), (R)- and (S)—, or (D)- and(L)-isomers may be prepared using chiral synthons or chiral reagents, orresolved using conventional techniques, for example, chromatography andfractional crystallization. Conventional techniques for thepreparation/isolation of individual enantiomers include chiral synthesisfrom a suitable optically pure precursor or resolution of the racemate(or the racemate of a salt or derivative) using, for example, chiralhigh pressure liquid chromatography (HPLC). When the compounds describedherein contain olefinic double bonds or other centres of geometricasymmetry, and unless specified otherwise, it is intended that thecompounds include both E and Z geometric isomers. Likewise, alltautomeric forms of curcuphenol are also intended to be included.

“Prodrug” is meant to indicate a curcuphenol compound that may beconverted under physiological conditions or by solvolysis to abiologically active compound. Thus, the term “prodrug” refers to ametabolic precursor of curcuphenol that is pharmaceutically acceptable.A prodrug may be inactive when administered to a subject in needthereof, but is converted in vivo to an active compound. Prodrugs aretypically rapidly transformed in vivo to yield the parent compound, forexample, by hydrolysis in blood. The prodrug compound often offersadvantages of solubility, tissue compatibility or delayed release in amammalian organism (see, Bundgard, H., Design of Prodrugs (1985), pp.7-9, 21-24 (Elsevier, Amsterdam)). A discussion of prodrugs is providedin Higuchi, T., et al., A.C.S. Symposium Series, Vol. 14, and inBioreversible Carriers in Drug Design, Ed. Edward B. Roche, AmericanPharmaceutical Association and Pergamon Press, 1987.

The term “prodrug” is also meant to include any covalently bondedcarriers, which release the active compound in vivo when such prodrug isadministered to a mammalian subject. Prodrugs of curcuphenol may beprepared by modifying functional groups present in the compound in sucha way that the modifications are cleaved, either in routine manipulationor in vivo, to the parent compound. Prodrugs include curcuphenolcompounds wherein a hydroxy, amino or mercapto group is bonded to anygroup that, when the prodrug is administered to a mammalian subject,cleaves to form a free hydroxy, free amino or free mercapto group,respectively. Examples of prodrugs include, but are not limited to,acetate, formate and benzoate derivatives of alcohol or amidederivatives of amine functional groups in curcuphenol and the like.

Also included are in vivo metabolic products of curcuphenol compounds.Such products may result from, for example, the oxidation, reduction,hydrolysis, amidation, esterification, and the like of the administeredcompound, primarily due to enzymatic processes. Accordingly, theinvention includes curcuphenol compounds produced by a processcomprising administering a compound of this invention to a mammal for aperiod of time sufficient to yield a metabolic product thereof. Suchproducts are typically identified by administering a radiolabelledcompound in a detectable dose to an animal, such as rat, mouse, guineapig, monkey, or to human, allowing sufficient time for metabolism tooccur, and isolating its conversion products from the urine, blood orother biological samples.

The “purity” of any given curcuphenol compound or mixture of curcuphenolcompounds in a composition may be specifically defined. For instance,certain compositions may comprise a curcuphenol compound that is atleast 80, 85, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, or 100% pure,including all decimals in between, as measured, for example, by highpressure liquid chromatography (HPLC).

“Stable compound” and “stable structure” are meant to indicate acompound that is sufficiently robust to survive isolation to a usefuldegree of purity from a reaction mixture, and formulation into anefficacious therapeutic agent.

By “statistically significant,” it is meant that the result was unlikelyto have occurred by chance. Statistical significance can be determinedby any method known in the art. Commonly used measures of significanceinclude the p-value, which is the frequency or probability with whichthe observed event would occur, if the null hypothesis were true. If theobtained p-value is smaller than the significance level, then the nullhypothesis is rejected. In simple cases, the significance level isdefined at a p-value of 0.05 or less.

A “subject,” as used herein, includes any animal that exhibits asymptom, or is at risk for exhibiting a symptom, which can be treated ordiagnosed with a composition described herein. Suitable subjects(patients) include laboratory animals (such as mouse, rat, rabbit, orguinea pig), farm animals, and domestic animals or pets (such as a cator dog). Mammals including non-human primates and, preferably, humanpatients, are included.

“Substantially” or “essentially” means nearly totally or completely, forinstance, 95%, 96%, 97%, 98%, 99% or greater of some given quantity.

“Substantially free” refers to the nearly complete or complete absenceof a given quantity for instance, less than about 10%, 5%, 4%, 3%, 2%,1%, 0.5% or less of some given quantity. For example, certaincompositions may be “substantially free” of cell proteins, membranes,nucleic acids, endotoxins, or other contaminants.

A “stereoisomer” refers to a compound made up of the same atoms bondedby the same bonds but having different three-dimensional structures,which are not interchangeable. The present invention contemplatesvarious stereoisomers and mixtures thereof and includes “enantiomers”,which refers to two stereoisomers whose molecules are nonsuperimposeablemirror images of one another.

A “tautomer” refers to a proton shift from one atom of a molecule toanother atom of the same molecule. The present invention includestautomers of any said compounds.

A “therapeutically effective amount” or “effective amount” includes anamount of a curcuphenol compound which, when administered to a mammal,preferably a human, is sufficient to increase MHC-I surface expressionin one or more cells and/or treat any one or more other conditionsdescribed herein. The amount of a curcuphenol compound which constitutesa “therapeutically effective amount” will vary depending on thecompound, the condition and its severity, the manner of administration,and the age of the mammal to be treated, but can be determined routinelyby one of ordinary skill in the art having regard to his own knowledgeand to this disclosure.

“Treatment” or “treating,” as used herein, includes any desirable effecton the symptoms or pathology of a disease or condition, and may includeeven minimal changes or improvements in one or more measurable markersof the disease or condition being treated. “Treatment” or “treating”does not necessarily indicate complete eradication or cure of thedisease or condition, or associated symptoms thereof. The subjectreceiving this treatment is any subject in need thereof. Exemplarymarkers of clinical improvement will be apparent to persons skilled inthe art.

Methods and Pharmaceutical Compositions

The methods and compositions described herein utilize or comprise one ormore curcuphenol compounds. Examples of “curcuphenol compounds” includecurcuphenol and its enantiomers, stereoisomers, diastereomers, and otherstereoisomeric forms, racemates, tautomers, metabolites, and prodrugsthat can increase MHC-I surface expression in a cell. Also included arepharmaceutically acceptable salts of the foregoing, including acid andbase addition salts.

Curcuphenol is a sesquiterpene phenol isolated from different marinesponges belonging to the genus Didiscus (see El Sayed et al., J NatProd. 65:1547-53, 2002, incorporated by reference in its entirety). Itis also referred to as phenol,2-(1,5-dimethyl-4-hexenyl)-5-methyl-,(S)—; Phenol,2-[(1S)-1,5-dimethyl-4-hexenyl]-5-methyl-(9Cl);(+)-Curcuphenol; (S)-(+)-Curcuphenol; and (S)-Curcuphenol.

In certain embodiments, a curcuphenol compound has the following Formula(I-A):

In some embodiments, the curcuphenol compound has the following Formula(I-B):

In some embodiments, the curcuphenol compound has the following Formula(II), also referred to as PC-02-113:

In some embodiments, the curcuphenol compound has the following Formula(III), also referred to as PC-02-113:

In some embodiments, the curcuphenol compound has the following Formula(IV), also referred to as PC-02-116:

In some embodiments, the curcuphenol compound has the following Formula(V), also referred to as PC-02-123:

Specific embodiments employ (S)-(+)-curcuphenol or analogs thereof. Forinstance, certain embodiments include one or more of(S)-(+)-15-hydroxycurcuphenol, (S)-(+)-12-hydroxycurcuphenol,(S)-(+)-12,15-dihydroxycurcuphenol, (S)-(+)-15-hydroxycurcuphenol-12-al,(S)-(+)-12-carboxy-10,11-dihydrocurcuphenol,(S)-(+)-12-hydroxy-10,11-dihydrocurcuphenol,(S)-(+)-4-[1-(2-hydroxy-4-methyl)phenyl)]pentanoic acid (11),(S)-curcuphenol-1alpha-D-glucopyranoside, (S)-(+)-4-nitrocurcuphenol(14), (S)-(+)-2-nitrocurcuphenol, (S)-(+)-curcuphenol-1-O-isonicotinate(see El Sayed et al., 2002, supra). FIG. 1 shows the formulae of(S)-(+)-curcuphenol and various other curcuphenol compounds. FIGS. 2A-2Bshow the formulae of curcuphenol compounds produced by coupling ofcurcuphenol with various carboxylic acids. Additional examples ofcurcuphenol compounds is described, for example, in Gul et al., BiochimBiophys Acta. 1770:1513-1519, 2007; Ono et al., Chem. Pharm. Bull.49:1581-1585, 2001; and Plano et al., Chem Biodivers. 8:1098-1111, 2011,incorporated by reference in their entireties. FIG. 10 shows theformulae of exemplary curcuphenol compounds, including curcuphenol andanalogs thereof. Certain embodiments may employ mixtures of any of thecurcuphenol compounds described herein.

In some embodiments, the curcuphenol compound(s) are chemicallysynthesized. The synthesis of various curcuphenol compounds isdescribed, for example, in El Sayed et al., 2002, supra; Gul et al.,2007, supra; Ono et al., 2001, supra; and Plano et al., 2011, supra).

In certain embodiments, the curcuphenol compound(s) are obtained frommarine sponge extracts or terrestrial plant extracts, for example, ofthe genera Didiscus, Myrmekioderma, Epipolapsis, Pseudopterogorgia,Elvira, or Laisanthaea. Exemplary species of these marine sponges andterrestrial plants include Didiscus oxeata, Myrmekioderma styx,Pseudopterogorgia rigida, Elvira biflora, and Laisanthaea podocephala.Certain aspects thus include the administration of marine sponge orterrestrial plant extract(s) that comprise one or more curcuphenolcompounds. In some instances, the purity of the curcuphenol compound(s)in the extract is about or at least about 50%, 60%, 70%, 80%, 90%, 95%,96%, 97%, 98%, 99%, or 100%.

Certain embodiments employ curcuphenol compounds or compositionscomprising the same to increase MHC-I expression in a cell, and therebyincrease the immunogenicity of the cell—that is, the ability of theimmune system (e.g., via interaction with CTLs) to target the cell fordestruction. Some embodiments therefore relate to method for increasingmajor histocompatibility complex class I (MHC-I) surface expression in acell, comprising contacting the cell with one or more curcuphenolcompounds or a composition that comprises the same. In some aspects,MHC-I surface expression is increased by about or at least about 10%,20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 100%, 150%, 200%, 300%, 400%,500%, 600%, 700%, 800%, 900%, or 1000% or more relative to an untreatedcontrol cell.

In some aspects, the curcuphenol compound(s) increase MHC-I surfaceexpression by increasing the expression of Transporter associated withAntigen Processing 1 (TAP-1), a transporter protein of the MHC-I antigenpresentation pathway. Hence, in certain aspects, the expression of TAP-1is increased by about or at least about 10%, 20%, 30%, 40%, 50%, 60%,70%, 80%, 90%, 100%, 150%, 200%, 300%, 400%, 500%, 600%, 700%, 800%,900%, or 1000% or more relative to an untreated control cell.

In certain aspects, the cell is a (diseased) cell characterized byreduced MHC-I surface expression (in its untreated state) relative to anon-diseased or otherwise normal or healthy cell of the same cell type.In some aspects, reduced MHC-I surface expression in the diseased cellis associated with or caused by reduced TAP-1 expression. Hence, in someaspects, the cell is a (diseased) cell characterized by reduced TAP-1expression (in its untreated state) relative to a non-diseased orotherwise normal or healthy cell of the same cell type.

In some aspects, after contacting with one or more curcuphenolcompounds, MHC-I surface expression and/or TAP-1 expression in thetreated cell is increased to a level that is comparable to the MHC-Isurface expression and/or TAP-1 expression of an otherwise normal orhealthy cell of the same cell type. For instance, in these and relatedaspects, MHC-I surface expression and/or TAP-1 expression can beincreased to about or within about 50%, 40%, 30%, 20%, 10%, or 5% of thelevels of MHC-I surface expression of the otherwise normal or healthycell of the same cell type.

In certain embodiments, the cell is a cancer or cancerous cell. Inspecific embodiments, the cancer cell is a metastatic or invasive cancercell. Examples of cancer cells include breast cancer cell, a cervicalcancer cell, a prostate cancer cell, a gastrointestinal cancer cell, alung cancer cell, an ovarian cancer cell, a testicular cancer cell, ahead and neck cancer cell, a bladder cancer cell, a kidney cancer cell(e.g., renal cell carcinoma), a squamous cell carcinoma, a CNS or braincancer cell, a melanoma cell, a non-melanoma cancer cell, a thyroidcancer cell, a endometrial cancer cell, an epithelial tumor cell, a bonecancer cell, or a hematopoietic cancer cell.

Examples or primary bone cancer cells include osteosarcomas,chondrosarcomas, and cells of the Ewing Sarcoma Family of Tumors(ESFTs). Examples of gastrointestinal cancer cells include esophagealcancer cells, stomach (gastric) cancer cell, pancreatic cancer cells,liver cancer cells, gallbladder (biliary) cancer cells, small intestinalcancer cells, colorectal cancer cells, anal or rectal cancer cells, andgastrointestinal carcinoid or stromal tumors.

Examples of lung cancer cells include adenocarcinomas, squamous-celllung carcinomas, small-cell lung carcinomas, and large-cell lungcarcinomas.

Particular examples of CNS or brain cancer cells include gliomas,meningiomas, pituitary adenomas, vestibular schwannomas, primary CNSlymphomas, neuroblastomas, and primitive neuroectodermal tumors(medulloblastomas). In some embodiments, the glioma is an astrocytoma,oligodendroglioma, ependymoma, or a choroid plexus papilloma. In someaspects, the brain cancer cell is a glioblastoma multiforme. In someembodiments, the glioblastoma multiforme is a giant cell glioblastoma ora gliosarcoma. In particular embodiments, the cancer cell is ametastatic cancer of the CNS, for instance, a cancer cell that hasmetastasized to the brain. Examples of such cancer cells include,without limitation, metastatic breast cancer cells, metastatic lungcancer cells, metastatic genitourinary tract cancer cells, metastaticgastrointestinal tract cancer cells (e.g., colorectal cancer cells,pancreatic carcinomas), osteosarcomas, melanomas, metastatic head andneck cancer cells, metastatic prostate cancer cells (e.g., prostaticadenocarcinomas), and metastatic lymphomas.

Examples of melanoma cells include those derived from lentigo maligna,lentigo maligna melanomas, superficial spreading melanomas, acrallentiginous melanomas, mucosal melanomas, nodular melanomas, polypoidmelanomas, desmoplastic melanomas, amelanotic melanomas, soft-tissuemelanomas, and uveal melanomas.

Examples of hematopoietic cancer cells include lymphoma cells, leukemiacells, and multiple myeloma cells. In some instances, the lymphoma cellis a T-cell lymphoma, B-cell lymphoma, small lymphocytic lymphoma,mangle cell lymphoma, anaplastic large cell lymphoma (ALCL), follicularlymphoma, Hodgkin's lymphoma, or non-Hodgkin's lymphoma. In particularinstances, the leukemia cell is chronic lymphocytic leukemia (CLL),hairy cell leukemia, acute lymphoblastic leukemia, myelocytic leukemia,acute myeloid or myelogenous leukemia, or chronic myelogenous leukemia.

In certain embodiments, the cell is in vitro, for example, in tissueculture. Methods of culturing cells including cancer or transformedcells are well-known in the art (see, for example, Animal Cell Culture(R. Freshney, ed., 1986); Freshney, R. I. (2005) Culture of AnimalCells, a Manual of Basic Technique, 5^(th) Ed. Hoboken N.J., John Wiley& Sons).

In certain embodiments, the cell is in a subject, and the methodcomprises administering the curcuphenol compound or related compositionto the subject. For the purposes of administration, the curcuphenolcompounds may be administered to a patient or subject as a raw chemicalor may be formulated as pharmaceutical compositions. Pharmaceuticalcompositions generally comprise a curcuphenol compound and apharmaceutically acceptable carrier, diluent, or excipient. Thecurcuphenol compound is typically present in the composition in anamount which is effective to treat a particular disease or condition ofinterest, as described herein, and preferably with acceptable toxicityto the subject. The activity of compound(s) can be determined by oneskilled in the art, for example, as described in the Examples below.Appropriate concentrations and dosages can be readily determined by oneskilled in the art.

A curcuphenol compound or related composition may be used in a methodfor treating essentially any disease or other condition in a subjectwhich would benefit from increased surface expression of MHC-Imolecules. In particular embodiments, the subject has cancer, and themethod comprises treating the cancer by administering one or morecurcuphenol compounds to the subject. As illustrated in FIG. 5, TAP-1 isfrequently down-regulated in cancer cell lines and surgically removedtumor cells. Without wishing to be bound by any one theory, it isbelieved that down-regulation of TAP-1 associates with reduced MHC-Isurface expression and thus reduced CTL-mediated kill of cancer cells.Restoration of MHC-I, for instance, via restoration of TAP-1 expression,has been shown to increase CTL-mediated killing of cancer cells in vitroand suppression of tumor growth in vivo (see, for example, Zhang et al.,2007, Lou et al., 2007, Lou et al., 2005, Alimonti et al., 2000).

Accordingly, in some aspects, the cancer is characterized by cancercells having reduced MHC-I surface expression (in an untreated state)and optionally reduced TAP-1 expression (in an untreated state) relativeto a non-cancerous cell of the same cell type. In particular aspects,the cancer is characterized by cancer cells having about or less thanabout 80%, 70%, 60%, 50%, 40%, 30%, 20%, 10%, 5% of the levels of MHC-Isurface expression and optionally TAP-1 expression of a non-cancerouscell of the same cell type.

In some aspects, the subject has or is at risk for having a metastaticor invasive cancer. As illustrated in FIG. 6, TAP-1 down-regulationstrongly correlates with disease progression and metastasis. Thus, insome aspects, the metastatic cancer is characterized by cancer cellshaving reduced MHC-I surface expression of MHC-I (in an untreated state)and optionally reduced TAP-1 expression (in an untreated state) relativeto a non-cancerous cell of the same cell type, or relative to anon-metastatic cancerous cell of the same cell type. In particularaspects, the cancer is characterized by metastatic cancer cells havingabout or less than about 80%, 70%, 60%, 50%, 40%, 30%, 20%, 10%, 5% ofthe levels of MHC-I surface expression and optionally TAP-1 expressionof a non-cancerous cell of the same cell type, or of a non-metastaticcancerous cell of the same cell type.

In some aspects, administration of the curcuphenol compound increasesMHC-I surface expression and optionally TAP-1 expression in about or atleast about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% of thecancer cells(s) by about or at least about 10%, 20%, 30%, 40%, 50%, 60%,70%, 80%, 90%, or 100%, 150%, 200%, 300%, 400%, 500%, 600%, 700%, 800%,900%, or 1000% or more relative to that of a control cell or populationof control cells. In some instances, the control cell(s) are from anuntreated state, for example, prior to any treatment, or from one ormore earlier-treated states, for example, following a series ofadministrations or treatments.

In some instances, administration of the curcuphenol compound increasesMHC-I surface expression and optionally TAP-1 expression in about or atleast about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or 100% of thecancer cells(s) to levels comparable to (for example, about or withinabout 50%, 40%, 30%, 20%, 10%, or 5% of) a reference standard, forexample, a reference standard of the average MHC-I surface expression orTAP-1 expression in cells of the same type from healthy (non-cancerous)individuals.

In some embodiments, increased MHC-I surface expression and optionallyincreased TAP-1 expression increases the immunogenicity of the cancercells, and thereby increases the immune response against the cancercells. In some instances, the immune response is a cytotoxic Tlymphocyte (CTL)-mediated immune response, and can include, for example,CTL activation, clonal expansion, and increased CTL effector function.Examples of CTL effector functions include the release of release thecytotoxins perforin, granzymes, and granulysin, and increased expressionof the CTL surface protein FAS ligand (FasL). In some instances,increased MHC-I surface expression and optionally increased TAP-Iexpression in the cancer cell(s) increases the CTL-mediated destructionof the cancer cell(s). For solid tumors, administration of one or morecurcuphenol compounds can reduce tumor expansion or reduce tumor size,for instance, by about or at least about 10%, 20%, 30%, 40%, 50%, 60%,70%, 80%, 90%, or 100% relative to an untreated state or anearlier-treated stated.

In some embodiments, the subject has a cancer selected from one or moreof breast cancer, cervical cancer, prostate cancer, gastrointestinalcancer, lung cancer, ovarian cancer, testicular cancer, head and neckcancer, bladder cancer, kidney cancer (e.g., renal cell carcinoma), softtissue sarcoma, squamous cell carcinoma, CNS or brain cancer, melanoma,non-melanoma cancer, thyroid cancer, endometrial cancer, an epithelialtumor, bone cancer, or a hematopoietic cancer.

Examples of lung cancers include adenocarcinomas, squamous-cell lungcarcinomas, small-cell lung carcinomas, and large-cell lung carcinomas.

Examples or primary bone cancers include osteosarcoma, chondrosarcoma,and the Ewing Sarcoma Family of Tumors (ESFTs).

Examples of gastrointestinal cancers include esophageal cancer, stomach(gastric) cancer, pancreatic cancer, liver cancer, gallbladder (biliary)cancer, small intestinal cancer, colorectal cancer, anal or rectalcancer, and gastrointestinal carcinoid or stromal tumors.

Examples of CNS or brain cancers include primary brain cancers andmetastatic brain cancers. Particular examples of brain cancers includegliomas, meningiomas, pituitary adenomas, vestibular schwannomas,primary CNS lymphomas, neuroblastomas, and primitive neuroectodermaltumors (medulloblastomas). In some embodiments, the glioma is anastrocytoma, oligodendroglioma, ependymoma, or a choroid plexuspapilloma. In some aspects, the subject has a glioblastoma multiforme.In specific aspects, the glioblastoma multiforme is a giant cellglioblastoma or a gliosarcoma. In particular embodiments, the cancer isa metastatic cancer of the CNS, for instance, a cancer that hasmetastasized to the brain. Examples of such cancers include, withoutlimitation, breast cancers, lung cancers, genitourinary tract cancers,gastrointestinal tract cancers (e.g., colorectal cancers, pancreaticcarcinomas), osteosarcomas, melanomas, head and neck cancers, prostatecancers (e.g., prostatic adenocarcinomas), and lymphomas.

Examples of melanomas include lentigo maligna, lentigo maligna melanoma,superficial spreading melanoma, acral lentiginous melanoma, mucosalmelanoma, nodular melanoma, polypoid melanoma, desmoplastic melanoma,amelanotic melanoma, soft-tissue melanoma, and uveal melanoma.

Examples of hematopoietic cancers include lymphomas, leukemias, andmultiple myelomas. In some instances, the lymphoma is a T-cell lymphoma,B-cell lymphoma, small lymphocytic lymphoma, mangle cell lymphoma,anaplastic large cell lymphoma (ALCL), follicular lymphoma, Hodgkin'slymphoma, or non-Hodgkin's lymphoma. In particular instances, theleukemia is chronic lymphocytic leukemia (CLL), hairy cell leukemia,acute lymphoblastic leukemia, myelocytic leukemia, acute myeloid ormyelogenous leukemia, or chronic myelogenous leukemia.

The use of curcuphenol compounds for treating cancers can be combinedwith other therapeutic modalities. For example, one or more curcuphenolcompounds can be administered to a subject before, during, or afterother therapeutic interventions, including symptomatic care,radiotherapy, surgery, transplantation, hormone therapy, immunotherapy,photodynamic therapy, antibiotic therapy, and administration ofanti-cancer agents, including any combination thereof. Symptomatic careincludes administration of corticosteroids, to reduce cerebral edema,headaches, cognitive dysfunction, and emesis, and administration ofanti-convulsants, to reduce seizures. Radiotherapy includes whole-brainirradiation, fractionated radiotherapy, and radiosurgery, such asstereotactic radiosurgery, which can be further combined withtraditional surgery.

Examples of anti-cancer agents include small molecules and therapeuticantibodies, among others known in the art. In certain embodiments, thesmall molecule is a cytotoxic or chemotherapeutic or anti-angiogenicagent. Particular examples include alkylating agents, anti-metabolites,anthracyclines, anti-tumor antibiotics, platinums, type I topoisomeraseinhibitors, type II topoisomerase inhibitors, vinca alkaloids, andtaxanes. In certain embodiments, the small molecule is selected from oneor more of chlorambucil, cyclophosphamide, cilengitide, lomustine(CCNU), melphalan, procarbazine, thiotepa, carmustine (BCNU),enzastaurin, busulfan, daunorubicin, doxorubicin, gefitinib, erlotinibidarubicin, temozolomide, epirubicin, mitoxantrone, bleomycin,cisplatin, carboplatin, oxaliplatin, camptothecins, irinotecan,topotecan, amsacrine, etoposide, etoposide phosphate, teniposide,temsirolimus, everolimus, vincristine, vinblastine, vinorelbine,vindesine, CT52923, paclitaxel, imatinib, dasatinib, sorafenib,pazopanib, sunitnib, vatalanib, geftinib, erlotinib, AEE-788,dichoroacetate, tamoxifen, fasudil, SB-681323, semaxanib, donepizil,galantamine, memantine, rivastigmine, tacrine, rasigiline, naltrexone,lubiprostone, safinamide, istradefylline, pimavanserin, pitolisant,isradipine, pridopidine (ACR16), tetrabenazine, bexarotene, glatirimeracetate, fingolimod, and mitoxantrone, including pharmaceuticallyacceptable salts and acids thereof.

In certain embodiments, the antibody is selected from one or more of3F8, 8H9, abagovomab, adecatumumab, afutuzumab, alacizumab (pegol),alemtuzumab, altumomab pentetate, amatuximab, anatumomab mafenotox,apolizumab, arcitumomab, bavituximab, bectumomab, belimumab,bevacizumab, bivatuzumab (mertansine), brentuximab vedotin, cantuzumab(mertansine), cantuzumab (ravtansine), capromab (pendetide), carlumab,catumaxomab, cetuximab, citatuzumab (bogatox), cixutumumab, clivatuzumab(tetraxetan), conatumumab, dacetuzumab, daclizumab, dalotuzumab,detumomab, drozitumab, ecromeximab, edrecolomab, elotuzumab,enavatuzumab, ensituximab, epratuzumab, ertumaxomab, etaracizumab,farletuzumab, FBTA05, figitumumab, flanvotumab, galiximab, gemtuzumab,ganitumab, gemtuzumab (ozogamicin), girentuximab, glembatumumab(vedotin), ibritumomab tiuxetan, icrucumab, igovomab, indatuximabravtansine, intetumumab, inotuzumab ozogamicin, ipilimumab (MDX-101),iratumumab, labetuzumab, lexatumumab, lintuzumab, lorvotuzumab(mertansine), lucatumumab, lumiliximab, mapatumumab, matuzumab,milatuzumab, mitumomab, mogamulizumab, moxetumomab (pasudotox),nacolomab (tafenatox), naptumomab (estafenatox), narnatumab,necitumumab, nimotuzumab, nivolumab, Neuradiab® (with or withoutradioactive iodine), NR-LU-10, ofatumumab, olaratumab, onartuzumab,oportuzumab (monatox), oregovomab, panitumumab, patritumab, pemtumomab,pertuzumab, pritumumab, racotumomab, radretumab, ramucirumab,rilotumumab, rituximab, robatumumab, samalizumab, sibrotuzumab,siltuximab, tabalumab, tanezumab, taplitumomab (paptox), tenatumomab,teprotumumab, TGN1412, ticilimumab, trastuzumab, tremelimumab,tigatuzumab, TNX-650, tositumomab, TRBS07, tucotuzumab (celmoleukin),ublituximab, urelumab, veltuzumab, volociximab, votumumab, andzalutumumab, including antigen-binding fragments thereof.

As noted above, certain embodiments include pharmaceutical compositionscomprising a curcuphenol compound as described herein and apharmaceutically acceptable carrier. Such pharmaceutical compositionscan also comprise one or more additional agents, as described herein,including, for example, anti-cancer agents.

Administration of the curcuphenol compounds, or their pharmaceuticallyacceptable salts, in pure form or in an appropriate pharmaceuticalcomposition, can be carried out via any of the accepted modes ofadministration of agents for serving similar utilities. Thepharmaceutical compositions can be prepared by combining a curcuphenolcompound with an appropriate pharmaceutically acceptable carrier,diluent or excipient, and may be formulated into preparations in solid,semi-solid, liquid or gaseous forms, such as tablets, capsules, powders,granules, ointments, solutions, suppositories, injections, inhalants,gels, microspheres, and aerosols.

Typical routes of administering such pharmaceutical compositionsinclude, without limitation, oral, topical, transdermal, inhalation,parenteral, sublingual, buccal, rectal, vaginal, and intranasal. Theterm parenteral as used herein includes subcutaneous injections,intravenous, intramuscular, intrasternal injection or infusiontechniques. Pharmaceutical compositions are formulated so as to allowthe active ingredients contained therein to be bioavailable uponadministration of the composition to a subject. Compositions that willbe administered to a subject or patient take the form of one or moredosage units, where for example, a tablet may be a single dosage unit,and a container of a compound of the invention in aerosol form may holda plurality of dosage units. Actual methods of preparing such dosageforms are known, or will be apparent, to those skilled in this art; forexample, see Remington: The Science and Practice of Pharmacy, 20thEdition (Philadelphia College of Pharmacy and Science, 2000). Thecomposition to be administered will, in any event, contain atherapeutically effective amount of a curcuphenol compound, or apharmaceutically acceptable salt thereof, for treatment of a disease orcondition of interest in accordance with the teachings herein.

A pharmaceutical composition may be in the form of a solid or liquid. Inone aspect, the carrier(s) are particulate, so that the compositionsare, for example, in tablet or powder form. The carrier(s) may beliquid, with the compositions being, for example, an oral syrup,injectable liquid or an aerosol, which is useful in, for example,inhalatory administration.

When intended for oral administration, the pharmaceutical composition ispreferably in either solid or liquid form, where semi-solid,semi-liquid, suspension and gel forms are included within the formsconsidered herein as either solid or liquid.

As a solid composition for oral administration, the pharmaceuticalcomposition may be formulated into a powder, granule, compressed tablet,pill, capsule, chewing gum, wafer or the like form. Such a solidcomposition will typically contain one or more inert diluents or ediblecarriers. In addition, one or more of the following may be present:binders such as carboxymethylcellulose, ethyl cellulose,microcrystalline cellulose, gum tragacanth or gelatin; excipients suchas starch, lactose or dextrins, disintegrating agents such as alginicacid, sodium alginate, Primogel, corn starch and the like; lubricantssuch as magnesium stearate or Sterotex; glidants such as colloidalsilicon dioxide; sweetening agents such as sucrose or saccharin; aflavoring agent such as peppermint, methyl salicylate or orangeflavoring; and a coloring agent.

When the pharmaceutical composition is in the form of a capsule, forexample, a gelatin capsule, it may contain, in addition to materials ofthe above type, a liquid carrier such as polyethylene glycol or oil.

The pharmaceutical composition may be in the form of a liquid, forexample, an elixir, syrup, solution, emulsion or suspension. The liquidmay be for oral administration or for delivery by injection, as twoexamples. When intended for oral administration, preferred compositioncontain, in addition to the present compounds, one or more of asweetening agent, preservatives, dye/colorant and flavor enhancer. In acomposition intended to be administered by injection, one or more of asurfactant, preservative, wetting agent, dispersing agent, suspendingagent, buffer, stabilizer and isotonic agent may be included.

The liquid pharmaceutical compositions, whether they be solutions,suspensions or other like form, may include one or more of the followingadjuvants: sterile diluents such as water for injection, salinesolution, preferably physiological saline, Ringer's solution, isotonicsodium chloride, fixed oils such as synthetic mono or diglycerides whichmay serve as the solvent or suspending medium, polyethylene glycols,glycerin, propylene glycol or other solvents; antibacterial agents suchas benzyl alcohol or methyl paraben; antioxidants such as ascorbic acidor sodium bisulfite; chelating agents such as ethylenediaminetetraaceticacid; buffers such as acetates, citrates or phosphates and agents forthe adjustment of tonicity such as sodium chloride or dextrose. Theparenteral preparation can be enclosed in ampoules, disposable syringesor multiple dose vials made of glass or plastic. Physiological saline isa preferred adjuvant. An injectable pharmaceutical composition ispreferably sterile.

A liquid pharmaceutical composition intended for either parenteral ororal administration should contain an amount of a curcuphenol compoundsuch that a suitable dosage will be obtained.

The pharmaceutical composition may be intended for topicaladministration, in which case the carrier may suitably comprise asolution, emulsion, ointment or gel base. The base, for example, maycomprise one or more of the following: petrolatum, lanolin, polyethyleneglycols, bee wax, mineral oil, diluents such as water and alcohol, andemulsifiers and stabilizers. Thickening agents may be present in apharmaceutical composition for topical administration. If intended fortransdermal administration, the composition may include a transdermalpatch or iontophoresis device.

The pharmaceutical composition may be intended for rectaladministration, in the form, for example, of a suppository, which willmelt in the rectum and release the drug. The composition for rectaladministration may contain an oleaginous base as a suitablenonirritating excipient. Such bases include, without limitation,lanolin, cocoa butter and polyethylene glycol.

The pharmaceutical composition may include various materials, whichmodify the physical form of a solid or liquid dosage unit. For example,the composition may include materials that form a coating shell aroundthe active ingredients. The materials that form the coating shell aretypically inert, and may be selected from, for example, sugar, shellac,and other enteric coating agents. Alternatively, the active ingredientsmay be encased in a gelatin capsule.

The pharmaceutical composition in solid or liquid form may include anagent that binds to the compound of the invention and thereby assists inthe delivery of the compound. Suitable agents that may act in thiscapacity include a monoclonal or polyclonal antibody, a protein or aliposome.

The pharmaceutical composition may consist of dosage units that can beadministered as an aerosol. The term aerosol is used to denote a varietyof systems ranging from those of colloidal nature to systems consistingof pressurized packages. Delivery may be by a liquefied or compressedgas or by a suitable pump system that dispenses the active ingredients.

Aerosols of compounds of the invention may be delivered in single phase,bi-phasic, or tri-phasic systems in order to deliver the activeingredient(s). Delivery of the aerosol includes the necessary container,activators, valves, subcontainers, and the like, which together may forma kit. One skilled in the art, without undue experimentation maydetermine preferred aerosols.

The pharmaceutical compositions may be prepared by methodology wellknown in the pharmaceutical art. For example, a pharmaceuticalcomposition intended to be administered by injection can be prepared bycombining a compound of the invention with sterile, distilled water soas to form a solution. A surfactant may be added to facilitate theformation of a homogeneous solution or suspension. Surfactants arecompounds that non-covalently interact with the compound of theinvention so as to facilitate dissolution or homogeneous suspension ofthe compound in the aqueous delivery system.

The curcuphenol compounds, or their pharmaceutically acceptable salts,are administered in a therapeutically effective amount, which will varydepending upon a variety of factors including the activity of thespecific compound employed; the metabolic stability and length of actionof the compound; the age, body weight, general health, sex, and diet ofthe patient; the mode and time of administration; the rate of excretion;the drug combination; the severity of the particular disorder orcondition; and the subject undergoing therapy.

In certain embodiments, a typical dosage of the curcuphenol compound maybe between about 0.2 mg per day and about 2 g per day, or between about1 mg and about 1 g per day, or between about 5 mg and about 500 mg, orbetween about 10 mg and about 250 mg per day, which is administered to asubject in need of treatment.

The frequency of administration of the compounds and compositionsdescribed herein may vary from once-a-day (QD) to twice-a-day (BID) orthrice-a-day (TID), etc., the precise frequency of administrationvarying with, for example, the patient's condition, the dosage, etc.

Curcuphenol compounds may also be administered simultaneously with,prior to, or after administration of one or more other therapeutic orbiologically active agents, dietary supplements, or any combinationthereof. Such combination therapy includes administration of a singlepharmaceutical dosage formulation which contains a curcuphenol compoundand one or more additional active agents, as well as administration ofthe curcuphenol compound and each active agent in its own separatepharmaceutical dosage formulation. For example, a curcuphenol compoundand the other active agent can be administered to the subject togetherin a single dosage composition, or each agent administered in separatedosage compositions. Where separate dosage compositions are used, thecurcuphenol compounds and one or more additional active agents can beadministered at essentially the same time, i.e., concurrently, or atseparately staggered times, i.e., sequentially; combination therapy isunderstood to include all these regimens.

It is understood that in the present description, combinations ofsubstituents and/or variables of the depicted formulae are permissibleonly if such contributions result in stable or reasonably stablecompounds.

It will also be appreciated by those skilled in the art that in theprocess described herein the functional groups of intermediate compoundsmay need to be protected by suitable protecting groups. Such functionalgroups include hydroxy, amino, mercapto, and carboxylic acid. Suitableprotecting groups for hydroxy include trialkylsilyl or diarylalkylsilyl(for example, t-butyldimethylsilyl, t-butyldiphenylsilyl ortrimethylsilyl), tetrahydropyranyl, benzyl, and the like. Suitableprotecting groups for amino, amidino and guanidino includet-butoxycarbonyl, benzyloxycarbonyl, and the like. Suitable protectinggroups for mercapto include —C(O)—R″ (where R″ is alkyl, aryl orarylalkyl), p-methoxybenzyl, trityl and the like. Suitable protectinggroups for carboxylic acid include alkyl, aryl or arylalkyl esters.Protecting groups may be added or removed in accordance with standardtechniques, which are known to one skilled in the art and as describedherein. The use of protecting groups is described in detail in Green, T.W. and P. G. M. Wutz, Protective Groups in Organic Synthesis (1999), 3rdEd., Wiley. As one of skill in the art would appreciate, the protectinggroup may also be a polymer resin such as a Wang resin, Rink resin or a2-chlorotrityl-chloride resin.

It will also be appreciated by those skilled in the art, although suchprotected derivatives of curcuphenol compounds may not possesspharmacological activity as such, they may be administered to a mammaland thereafter metabolized in the body to form compounds of theinvention which are pharmacologically active. Such derivatives maytherefore be described as “prodrugs”. All prodrugs of compounds of thisinvention are included within the scope of the invention.

Furthermore, all curcuphenol compounds which exist in free base or acidform can be converted to their pharmaceutically acceptable salts bytreatment with the appropriate inorganic or organic base or acid bymethods known to one skilled in the art. Salts of the compounds of theinvention can be converted to their free base or acid form by standardtechniques.

EXAMPLES Example 1 Curcuphenol-Containing Extracts Induce TAP-1 PromoterExpression and MHC-I Surface Expression

Experiments were performed to identify marine sponge extracts thatinduce TAP-1 promoter expression and MHC-I surface expression in LMD:TAPreporter cells. LMD:TAP reporter cells were derived from metastaticTAP-1 and MHC class 1-deficient murine prostate cancer cells andmodified to contain a vector that expresses enhanced green fluorescentprotein (EGFP) under the control of the TAP-1 promoter.

LMD:TAP reporter cells were exposed to marine sponge extracts. Flowcytometry was used to measure expression at the TAP-1 promoter (EGFPexpression) and cell surface levels of MHC-I (MAb staining). As shown inFIGS. 7A and 7B, extract 1 increased surface expression of MHC-I (7A)and expression at the TAP-1 promoter (7B, as indicated by EGFP). Asshown in FIGS. 8A and 8B, extract 3 increased surface expression ofMHC-I (8A) and expression at the TAP-1 promoter (8B, as indicated byEGFP). As shown in FIGS. 9A and 9B, extract 5 also increased surfaceexpression of MHC-I (9A) and expression at the TAP-1 promoter (9B, asindicated by EGFP). Similar results were shown for extract 2.

Extracts 1-3 and 5 were further fractionated into purified extracts andtested by serial dilution to identify the compounds and concentrationswith the highest percent activity and the lowest toxicity. Therelatively pure extract identified as fraction 1A-b (at 0.018 mg/mL)displayed significant ability to increase both TAP-1 promoter activityand MHC-I expression (110% activity relative to IFN-γ positive control;% activity=x−μn/μp−μn, where x is the average GFP fluorescenceintensity, and μn and μp represent the average of the DMSO negativecontrol and IFN-γ positive control), and minimal toxicity (cell numbersup to 20% below the average cell number of the negative DMSO controlminus 3 standard deviation). The active compound in this extract wasidentified as curcuphenol.

Example 2 Curcuphenol and Analogs Induce MHC-I Expression in Lung TumorCells

Curcuphenol and four curcuphenol analogs (see FIG. 10) were synthesizedand tested for the ability to increase MHC-I expression in TC1/A9 murinelung tumor cells. Toxicity was also measured by propidium iodide (PI)exclusion.

The results are shown in FIGS. 11A and 11B. FIG. 11A shows thatcurcuphenol and the analog PC-02/113 had the best activity bysignificantly increasing MHC-I expression relative to controls. FIG. 11Bshows that these compounds also had acceptable levels of toxicity, withPC-02/113 showing the best combination of activity and toxicity.

Example 3 Testing of Curcuphenol Compound in Mouse Tumor Model

The compound PC-02-113 is selected for further testing in animalstudies. This compound is tested for its Maximum Tolerable Dose (MTD) inmice. Single intraperitoneal (IP) doses are administered as shown inTable E1 below, followed by clinical observation for 14 days.

TABLE E1 Maximum Tolerable Dose Study Plan # Mice Agent Dose (C57BI/6)Curcuphenol, PC-02-113 0.05 mg/kg 3 (10X less) Curcuphenol, PC-02-113 0.5 mg/kg 3 Curcuphenol, PC-02-113  5.0 mg/kg 3 (10X more) Vehiclealone n/a 3

This compound is also tested for its anti-tumor effects in mice injectedwith A9 tumor cells as shown in Table E2 below.

TABLE E2 Anti-Tumor Effect Study Plan # Mice Agent Dose (C57BI/6) [A9 +vect] n/a 8 [A9 + vect] + As determined 8 [Curcuphenol, PC-02-113] byMTD [A9 + vect] + [TSA] 0.5 mg/kg 8

The curcuphenol compound is administered 3 times a week (M, W, F) forfour weeks by intraperitoneal (IP) injection. Tumors are measured 3times a week: tumors from 4 mice/group are collected forimmunohistochemistry (IHC) analysis, and tumors from the other 4mice/group are collected for analysis of tumor-infiltrating lymphocytes(TIL). Lymph nodes, liver, brain, adrenal glands, spleen and blood arecollected from mice and analyzed at the end of the study.

1-40. (canceled)
 41. A method for treating cancer, comprisingadministering a curcuphenol compound to a subject having cancer, whereinsaid curcuphenol compound is

or a pharmaceutically acceptable salt thereof; and wherein said canceris carcinoma.
 42. The method of claim 41, wherein the cancer ischaracterized by cancer cells in an untreated state having reduced MHC-1surface expression and optionally reduced TAP-1 expression relative tonon-cancerous cells of the same cell type.
 43. The method of claim 42,wherein MHC-1 surface expression and optionally TAP-1 expression in thecancer cell(s) is increased by at least about 10% relative to a controlcell wherein optionally increased MHC-1 surface expression andoptionally TAP-1 expression increases a CTL-mediated immune responseagainst the cancer cells after administration of said curcuphenolcompound.
 44. The method of claim 41, wherein the cancer is carcinoma ofthe lung or prostate.
 45. The method of claim 41, further comprisingadministering an additional cancer therapy wherein optionally theadditional cancer therapy selected from one or more of an anti-canceragent, radiotherapy, surgery, transplantation, photodynamic therapy,symptomatic care, and antibiotic therapy wherein optionally theanti-cancer agent is selected from a small molecule and an antibodywherein optionally the small molecule is a cytotoxic, chemotherapeutic,or anti-angiogenic agent wherein optionally the small moleculecytotoxic, chemotherapeutic, or anti-angiogenic agent is selected fromone or more of alkylating agents, anti-metabolites, anthracyclines,anti-tumor antibiotics, platinums, type I topoisomerase inhibitors, typeII topoisomerase inhibitors, vinca alkaloids, and taxanes whereinoptionally the small molecule is selected from one or more ofchlorambucil, cyclophosphamide, cilengitide, lomustine, melphalan,procarbazine, thiotepa, carmustine (BCNU), enzastaurin, busulfan,daunorubicin, doxorubicin, gefitinib, erlotinib idarubicin,temozolomide, epirubicin, mitoxantrone, bleomycin, cisplatin,carboplatin, oxaliplatin, camptothecins, irinotecan, topotecan,amsacrine, etoposide, etoposide phosphate, teniposide, temsirolimus,everolimus, vincristine, vinblastine, vinorelbine, vindesine, CT52923,paclitaxel, imatinib, dasatinib, sorafenib, pazopanib, sunitnib,vatalanib, geftinib, erlotinib, AEE-788, dichoroacetate, tamoxifen,fasudil, SB-681323, semaxanib, donepizil, galantamine, memantine,rivastigmine, tacrine, rasigiline, naltrexone, lubiprostone, safinamide,istradefylline, pimavanserin, pitolisant, isradipine, pridopidine,tetrabenazine, bexarotene, glatirimer acetate, fingolimod, andmitoxantrone, including pharmaceutically acceptable salts and acidsthereof. where optionally the antibody is selected from one or more of3F8, 8H9, abagovomab, adecatumumab, afutuzumab, alacizumab, alemtuzumab,altumomab pentetate, amatuximab, anatumomab mafenotox, apolizumab,arcitumomab, bavituximab, bectumomab, belimumab, bevacizumab,bivatuzumab (mertansine), brentuximab vedotin, cantuzumab, cantuzumab,capromab, carlumab, catumaxomab, cetuximab, citatuzumab, cixutumumab,clivatuzumab, conatumumab, dacetuzumab, daclizumab, dalotuzumab,detumomab, drozitumab, ecromeximab, edrecolomab, elotuzumab,enavatuzumab, ensituximab, epratuzumab, ertumaxomab, etaracizumab,farletuzumab, FBTA05, figitumumab, flanvotumab, galiximab, gemtuzumab,ganitumab, gemtuzumab, girentuximab, glembatumumab, ibritumomabtiuxetan, icrucumab, igovomab, indatuximab ravtansine, intetumumab,inotuzumab ozogamicin, ipilimumab, iratumumab, labetuzumab, lexatumumab,lintuzumab, lorvotuzumab, lucatumumab, lumiliximab, mapatumumab,matuzumab, milatuzumab, mitumomab, mogamulizumab, moxetumomab(pasudotox), nacolomab, naptumomab, narnatumab, necitumumab,nimotuzumab, nivolumab, Iodine 1-131 monoclonal antibody 81c6, NR-LU-10,ofatumumab, olaratumab, onartuzumab, oportuzumab (monatox), oregovomab,panitumumab, patritumab, pemtumomab, pertuzumab, pritumumab,racotumomab, radretumab, ramucirumab, rilotumumab, rituximab,robatumumab, samalizumab, sibrotuzumab, siltuximab, tabalumab,tanezumab, taplitumomab, tenatumomab, teprotumumab, TGN1412,ticilimumab, trastuzumab, tremelimumab, tigatuzumab, TNX-650,tositumomab, TRBS07, tucotuzumab, ublituximab, urelumab, veltuzumab,volociximab, votumumab, and zalutumumab, including antigen-bindingfragments thereof.